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Objective: The clinical course of human metapneumovirus (hMPV) infection is similar to that of coronavirus 2019 disease (COVID-19). However, community-acquired hMPV infections in adults have not yet been sufficiently investigated. We examined the detection status of hMPV antigens and the clinical features of positive patients during the first wave of COVID-19, which coincided with the epidemic season of hMPV infection in Japan. Methods: In this cross-sectional, observational, and single-center study, we recruited consecutive individuals who visited the Japan Agricultural Cooperatives Kochi Hospital due to fever, respiratory symptoms, or close contact with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infected persons during the period from January to May 2020. Results: The positive rate of immunochromatography for hMPV antigens from nasopharyngeal swabs was 9.5% (4/42), and four positive cases were community-acquired pneumonia (CAP) (5.3% of all CAP). The positive rate of hMPV antigens in the CAP group (30.8%, 4/13) was higher than that in the non-pneumonia group (0.0%, 0/19) (p < 0.05). The average age of the four adult patients with CAP was 69.8 years (range 35–93). Mean white blood cell counts and C-reactive protein blood levels were 6,250 cells/μL (3,500–12,180) and 4.30 mg/dL (4.05–7.04), respectively. Chest computed tomography images were diverse and two patients showed dense consolidation. No multi-organ disorder was noted during the clinical course in any of the four cases, and their prognoses were good. Conclusion: hMPV infection may be considered in the differential diagnosis of COVID-19 and CAP in Japan under the preventive measures for SARS-CoV-2 infection, at least during the epidemic season of hMPV infection.

Chest symptoms and pleural effusion due to serositis in familial Mediterranean fever (FMF) are occasionally misdiagnosed as acute pneumonia. However, the actual pulmonary involvement of FMF is extremely rare. A 67-year-old man was referred to our hospital due to repeated and transient anterior chest pain. Chest images revealed a moderate amount of pericardial fluid, slight bilateral pleural effusion, and infiltrations in both lower lung lobes. Colchicine treatment without antibiotics rapidly improved these symptoms and findings. Pericarditis, pleurisy and the response to colchicine indicated FMF. FMF should be considered as a causative disease of pulmonary infiltrations, especially if it occurs repeatedly.

The incidence rate of pulmonary nontuberculous mycobacterial disease (PNTMD) in Japan is the highest among major industrialized nations. Although the typical clinical course and radiological manifestations of PNTMD are different from those of pulmonary tuberculosis (TB), confusion about these mycobacterial diseases leads to a diagnostic pitfall. Diagnostic challenges include the coexistence of Mycobacterium tuberculosis (MTB) and nontuberculous mycobacteria (NTM), false positives for NTM in MTB nucleic acid amplification tests, microbial substitution, and abnormal radiological manifestations caused by NTM. Features of extrapulmonary NTM diseases, such as pleurisy, vertebral osteomyelitis, and disseminated disease, are different from the corresponding tuberculous diseases. Moreover, the immunological background of the patient (status of human immunodeficiency virus infection with or without antiviral therapy, continuation or discontinuation of immunosuppressive therapy, use of immune checkpoint inhibitor, pregnancy and delivery, etc.) influences the pathophysiology of mycobacterial diseases. This review describes the varying clinical presentations of NTM disease with emphasis on the differences from TB. J. Med. Invest. 68 : 220-227, August, 2021.

Esophageal cancer is one of the most frequent causes of cancer-related deaths worldwide. This is due to its asymptomatic nature or mild nonspecific symptoms. Most patients are diagnosed after appearance of prominent symptoms, and tumors are frequently accompanied by severe infiltration. Therefore, molecular biomarkers for the prognosis of early-stage esophageal cancer are desired. In this study, we examined the prognostic potential of lipase H (LIPH), a recently reported biomarker for lung adenocarcinoma and squamous carcinoma. We found that LIPH mRNA is also frequently upregulated in esophageal adenocarcinoma. Immunohistochemical analysis confirmed LIPH protein expression in various esophageal tumor tissue sections. Interestingly, higher expression of LIPH in esophageal adenocarcinoma showed a positive correlation with longer survival of patients. Our data suggest that LIPH may have prognostic value for esophageal cancer.

Lung cancer is one of the most frequent causes of cancer-related death worldwide. However, molecular markers for lung cancer have not been well established. To identify novel genes related to lung cancer development, we surveyed publicly available DNA microarray data on lung cancer tissues. We identified lipase member H (LIPH, also known as mPA-PLA1) as one of the significantly upregulated genes in lung adenocarcinoma. LIPH was expressed in several adenocarcinoma cell lines when they were analyzed by quantitative real-time polymerase chain reaction (qPCR), western blotting, and sandwich enzyme-linked immunosorbent assay (ELISA). Immunohistochemical analysis detected LIPH expression in most of the adenocarcinomas and bronchioloalveolar carcinomas tissue sections obtained from lung cancer patients. LIPH expression was also observed less frequently in the squamous lung cancer tissue samples. Furthermore, LIPH protein was upregulated in the serum of early- and late-phase lung cancer patients when they were analyzed by ELISA. Interestingly, high serum level of LIPH was correlated with better survival in early phase lung cancer patients after surgery. Thus, LIPH may be a novel molecular biomarker for lung cancer, especially for adenocarcinoma and bronchioloalveolar carcinoma.

Expression of chemokine receptors by tumors, specifically CCR4 on cutaneous T cell lymphomas, is often associated with a poor disease outcome. To test the hypothesis that chemokine receptor-expressing tumors can be successfully controlled by delivering toxins through their chemokine receptors, we have generated fusion proteins designated chemotoxins: chemokines fused with toxic moieties that are nontoxic unless delivered into the cell cytosol. We demonstrate that chemokines fused with human RNase eosinophil-derived neurotoxin or with a truncated fragment of Pseudomonas exotoxin 38 are able to specifically kill tumors in vitro upon internalization through their respective chemokine receptors. Moreover, treatment with the thymus and activation-regulated chemokine (CCL17)-expressing chemotoxin efficiently eradicated CCR4-expressing cutaneous T cell lymphoma/leukemia established in NOD-SCID mice. Taken together, this work represents a novel concept that may allow control of growth and dissemination of tumors that use chemokine receptors to metastasize and circumvent immunosurveillance.

Despite its potency, the wider use of immunotherapy for B cell malignancies is hampered by the lack of well-defined tumor-specific Ags. In this study, we demonstrate that an evolutionarily conserved 37-kDa immature laminin receptor protein (OFA-iLRP), a nonimmunogenic embryonic Ag expressed by a variety of tumors, is rendered immunogenic if targeted to the APCs using the CCR6 ligands MIP3α/CCL20 and mDF2β. The CCR6 targeting facilitated efficient Ag cross-presentation and induction of tumor-neutralizing CTLs. Although the Ag targeting alone, without activation of dendritic cells (DCs), is proposed to induce tolerance, and MIP3α does not directly activate DCs, the MIP3α-based vaccine efficiently induced protective and therapeutic antitumor responses. The responses were as strong as those elicited by the OFA-iLRP fusions with moieties that activated DCs and Th1-type cytokine responses, mDF2β, or mycobacterial Hsp70 Ag. Although the same cDNA encodes the dimerized high-affinity mature 67-kDa mLRP that is expressed in normal tissues to stabilize the binding of laminin to cell surface integrins, the vaccines expressing OFA-iLRP elicited long-term protective CD8+ T cell-mediated memory responses against syngeneic B cell lymphoma, indicating the potential application of these simple vaccines as preventive and therapeutic formulations for human use.

Non-small cell lung cancer patient was found to have auto-antibodies against L-Myc in both the serum and pleural effusion. The titer of anti-L-Myc antibodies was higher in the pleural effusion than in the serum. Overexpression of L-Myc without DNA amplification was observed in the tumor cells. L-Myc antigen was not detected in either the serum or the pleural effusion. Anti-nuclear antibodies were also detected in both the serum and pleural effusion, although this patient did not have collagen-vascular disease.

Regulatory CD25+CD4+ T cells (Tregs) play an important role in the control of peripheral tolerance. In this study we demonstrate that human peripheral blood Tregs can be divided into two distinct populations based on the expression of CCR4. The majority (∼75%) of freshly isolated Tregs express CCR4 and presumably represent memory-type Tregs. Interestingly, CCR4− Tregs require anti-CD3 Ab-mediated activation to acquire a regulatory activity, while CCR4+ Tregs appear to be already primed to suppress the proliferation of CD8+ T cells. CCR4 is also expressed on CD25lowCD4+ T cells (CCR4+ non-Tregs) that mostly suppress Th1-type polarization without affecting T cell proliferation, presumably via the production of immunomodulatory cytokines like IL-10. In contrast, CCR4+ Tregs express FasL to primarily regulate T cell proliferation via a contact-mediated process involving FasL/Fas signaling, a major regulatory pathway of T cell homeostasis. Finally, we also demonstrate that the depletion of CCR4+ T cells leads to Th1-type polarization of CD4+ T cells and augmentation of CD8+ T cell responses to tumor Ags.

A 55-year-old woman visited our hospital for further examination of abnormal shadows on chest radiographs. Her routine chest radiograph showed two nodular shadows in the right lower lung field. A chest CT scan revealed other nodules, small patchy shadows in both lung fields, and enlargement of the mediastinal lymphnodes (#2, 3). Laboratory data showed polyclonal hyperimmunoglobulinemia. This case was initially considered on the basis of a transbronchial lung biopsy to be a plasma cell granuloma. However, serum gammaglobulin levels gradually increased, and video-assisted thoracoscopic surgery was performed to aid in making a definite diagnosis. Biopsy specimens revealed lymphoid follicles with plasma cells which were stained with both anti-kappa chain and anti-lambda chain antibodies. The patient was treated with prednisolone (50 mg/day), and the serum gammaglobulin level and the shadows on the chest CT were temporarily slightly improved. During the clinical course, her laboratory data and histological specimens were re-examined, and the final diagnosis was multicentric Castleman's disease.

TSC-36 (TGF-β1-stimulated clone 36) is a TGF-β1 inducible gene whose product is an extracellular glycoprotein that contains a single follistatin module. TSC-36 is highly expressed in the lung, but its physiological function is unknown. In an attempt to elucidate it, we investigated the effect of TSC-36 on proliferation of human lung cancer cell lines. We found a correlation between expression of TSC-36 and cell growth: TSC-36 mRNA was not detected in cells derived from small cell lung cancer (SCLC) cells, a highly aggressive neoplasm, but was detected in some non-small cell lung cancer (NSCLC) cells, a moderately aggressive neoplasm. This suggested an antiproliferative function for TSC-36. To address this question, NSCLC PC-14 cells, which express very low level of TSC-36 protein, were transfected with TSC-36 cDNA and the proliferative capacity of stable transfectants was determined by measuring the doubling time, colony forming activity in soft agar and the level of incorporation of 3 H-thymidine into DNA. Under normal culture conditions, the transfected cells showed a longer doubling time, lower plating efficiency and lower rate of DNA synthesis than the parental cells and the control neo transfectant cells. These findings suggested that expression of TSC-36 caused growth inhibition in human lung cancer cells.

The p16INK4A tumor suppressor gene is frequently inactivated in non-small cell lung carcinoma (NSCLC) and is less frequently inactivated in small cell lung carcinoma (SCLC) by mutation, deletion or DNA methylation. There are several reports that the reintroduction of the p16INK4A gene into p16(-) NSCLC cells results in significant growth suppression. However, there have been no reports of reintroduction of the p16INK4A gene into SCLC cells. To assess the biological significance of p16INK4A inactivation in the development of SCLC, full-length p16INK4A cDNA was introduced into an SCLC cell line, Ms-13, in which the p16INK4A protein was not detected. SCLC cells stably transfected with the p16INK4A expression vector formed only 2-16% of the number of neomycin-resistant colonies formed by cells transfected with a control vector, and no expression of exogenous p16INK4A protein was detected in any of 16 expanded clones. Transient transfection of the p16INK4A gene into SCLC cells resulted in exogeneous p16INK4A protein expression and dephosphorylation of endogenous retinoblastoma (RB) protein. These results suggest that the restoration of the p16INK4A function suppresses the growth of SCLC cells by dephosphorylation of the RB protein. Therefore, inactivation of p16INK4A may play an important role in the enhancement of growth of p16INK4A(-) RB(+) SCLC tumors in vivo.

We examined by immunoblotting antibodies that bind to p16INK4A tumor suppressor gene products in the sera of patients with lung cancer. Sera from 15 to 102 patients, including 6 with adenocarcinoma, 3 with squamous cell carcinoma, 2 with large cell carcinoma and 4 with small cell carcinoma, reacted with a glutathione-S-transferase (GST) fusion protein containing whole human anti-p16INK4A, produced by Escherichia coli. However, sera from 30 normal volunteers did not react with the GST-p16INK4A fusion protein. The background such as age, gender, performance status, histology, stage, smoking history, and prior treatment was not significantly different between the patients with and without anti-GST-p16INK4A antibodies. Circulating p16INK4A tumor suppressor products were not detected in any individuals with lung cancer or in the normal controls.

CCR4 is a chemokine receptor which is preferentially expressed on T helper 2 (Th2) CD4+ cells and CD25+CD4+ T regulatory cells (Tregs). Tregs and Th2-type cytokines (IL-10 and IL-4) were implicated in suppression of anti-tumor immune responses. We have hypothesized that depletion of CCR4+CD4+ cells, particularly Tregs and Th2 cells, may improve anti-tumor T cell immune responses. To test this, we generated a recombinant protein consisting of TARC (CCL17, ligand for CCR4) fused with 38kD truncated form of Pseudomonas exotoxin (PE38), designated TARC-PE38. TARC-PE38 efficiently and specifically killed CCR4+ cells, including sorted human peripheral blood CCR4+CD4+ and CCR4+ Tregs. We demonstrate that, while CCR4+non-Tregs mostly suppress Th1-type polarization without affecting T cell proliferation, CCR4+Tregs primarily regulate T cell proliferation via contact-mediated suppressive pathways. As a result, depletion of CCR4+ cells enhance proliferation of T cells and promote the Th1-type polarized responses. Moreover, the depletion also significantly enhanced tumor antigen-specific T cell responses in vitro. Importantly, in vivo depletion of CCR4+ cells augmented tumor - specific CD8+ CTL responses improving survival of tumor bearing mice, when combined with cancer DNA vaccines. Taken together, our data suggest that poor immune responses to tumors can be reversed by transiently depleting suppressive CCR4+ cells. This work was supported by the Intramural Research Program of the National Institute on Aging, NIH.